sq-23377 and leucine-methyl-ester

Recently, long-term preculture with IL-4 or IL-7 has been reported to induce IFN-gamma-producing ability in naive CD4+ T cells without stimulation via TCR. The mechanism of IFN-gamma-transcription in naive CD4+ T cells precultured with IL-4 was analyzed and compared with that in typical Th1 cells by focusing on the TATA proximal and first intronic regulatory regions of the IFN-gamma gene. Both regulatory regions in these IL-4-primed naive CD4+ T cells, which produce a large amount of IFN-gamma upon stimulation with PMA and ionomycin, were completely methylated in contrast to the same hypomethylated regions in Th1 cells. DNase I hypersensitive site analysis suggested that both regulatory regions in IL-4-primed naive CD4+ T cells were not active for IFN-gamma-expression. Moreover, we demonstrated that the composition of transcriptional factors that can bind to the proximal regulatory region is different between IL-4-primed naive CD4+ T cells and Th1 cells. These results indicated that the transcriptional machinery involved in the expression of the IFN-gamma gene by CD4+ T cells varied depending on their modes of differentiation in both the responsive regulatory regions and the specific nuclear factors.

Topics: CD4-Positive T-Lymphocytes; Cell Differentiation; Cells, Cultured; DNA; DNA Methylation; Fetal Blood; Gene Expression Regulation; Humans; Interferon-gamma; Interleukin-4; Introns; Ionomycin; Leucine; Recombinant Proteins; Regulatory Sequences, Nucleic Acid; TATA Box; Tetradecanoylphorbol Acetate; Th1 Cells; Transcription Factors; Transcription, Genetic

1. Uridine triphosphate (UTP), uridine diphosphate (UDP), cytidine triphosphate (CTP), and deoxythymidine triphosphate (TTP) caused concentration-dependent increases in the release of thromboxane A2 (TXA2) from cultured glia prepared from the newborn rat cerebral cortex. Although each of the pyrimidine nucleotides displayed similar potencies, CTP and TTP were considerably less effective than either UTP or UDP. The purine nucleotide ATP was equally as potent as the pyrimidine nucleotides but was marginally less effective than either UTP or UDP. 2. The ability of UTP, UDP, TTP, and CTP to promote TXA2 release from cultured glia was inhibited in a concentration-dependent manner by suramin and was markedly reduced when incubations were performed either in Ca(2+)-free medium or on cultures which had been maintained in serum-free growth medium for 4 days prior to experimentation. 3. Challenges with UTP and UDP in combination were found to elicit a response which was no different from the effects of these nucleotides alone; in addition, their effects were reversed by the phospholipase A2 inhibitor ONO-RS-082. A slight reduction in UTP- and UDP-stimulated TXA2 release was observed in cultures grown in the presence of leucine methyl ester, a treatment reported to limit microglial survival. 4. These results suggest that glia are targets for extracellular pyrimidine nucleotides and that their ability to release eicosanoids from these cells may be important in the brain's response to damage.

Topics: Aminobenzoates; Animals; Animals, Newborn; Calcium; Cells, Cultured; Cerebral Cortex; Chlorobenzoates; Cinnamates; Culture Media, Serum-Free; Cytidine Triphosphate; Drug Interactions; Ionomycin; Ionophores; Leucine; Neuroglia; ortho-Aminobenzoates; Phospholipases A; Phospholipases A2; Rats; Thromboxane A2; Thymine Nucleotides; Uridine Diphosphate; Uridine Triphosphate